Immunotherapy of Cancer Using Heat Shock Proteins

Frontiers in Bioscience : a Journal and Virtual Library. Jan, 2002  |  Pubmed ID: 11779704

Tumor derived heat shock protein (hsp)-peptide complexes (particularly hsp70 and grp94/gp96) have been demonstrated to serve as effective vaccines, producing anti-tumor immune responses in animals and in man. This approach utilizes the peptide binding properties of stress proteins which are responsible for their functions as molecular chaperones in numerous cellular processes. The present review briefly introduces the reader to the basic stress protein families, i.e. heat shock and glucose regulated proteins, their regulation, compartmentalization and family members. It then introduces the reader to aspects of hsps/grp function and interactions with the host's immune system. An overview of the conventional uses of hsp/grp vaccines as autologous vaccines derived from cancers is presented. We then discuss other stress protein related vaccination approaches. This includes the use of recombinant antigens, both proteins and peptides, naturally complexed to hsp/grps; hsp/grp DNA vaccines, hsp/grp fusion proteins and cell based hsp/grp vaccines. The advantages and disadvantages of each vaccination approach are discussed. Lastly, means of further enhancing the already potent activity of stress protein vaccines are presented, specifically the use of hyperthermia or CTLA-4 blockade as adjuvants.

Development of a Recombinant HSP110-HER-2/neu Vaccine Using the Chaperoning Properties of HSP110

Cancer Research. Mar, 2002  |  Pubmed ID: 11912148

Several studies have shown that when purified from a tumor, certain heat shock proteins (HSPs) can function as effective vaccines against the same tumor by virtue of their ability to bind tumor-specific peptides. However, only a small fraction of the associated peptides would be expected to be immunogenic, in addition to which, the clinical application of this vaccine requires the availability of a surgical specimen of sufficient quantity for purification of the HSP. The present study describes a new approach for the development of natural HSP vaccines that do not have these limitations. This approach uses a recombinant HSP that is noncovalently bound to a recombinant tumor protein antigen by heat shock. HSP110 has been selected for this purpose, because it has been shown to be a highly efficient molecular chaperone in binding to large protein substrates. We show that a "natural chaperone complex" between HSP110 and the intracellular domain (ICD) of human epidermal growth factor receptor 2 protein (HER-2)/neu is formed by heat shock. This HSP110-ICD vaccine elicited both CD8(+) and CD4(+) T-cell responses against ICD as determined by an antigen-specific IFN-gamma production in an enzyme-linked immunospot assay (ELISPOT). In vivo depletion studies revealed that the CD8(+) T-cell response was independent of CD4(+) T-cell help. The HSP110-ICD complex also significantly enhanced ICD-specific antibody responses relative to that seen with ICD alone. No CD8(+) T cell or antibody response was detected against HSP110. The use of recombinant HSP110 to form natural chaperone complexes with large protein antigens represents a new and powerful approach for the design of protein-targeted cancer vaccines.

Hsp110 Over-expression Increases the Immunogenicity of the Murine CT26 Colon Tumor

Cancer Immunology, Immunotherapy : CII. Aug, 2002  |  Pubmed ID: 12111119

Several studies have suggested a positive correlation between heat shock protein (hsp) expression and tumor immunogenicity. Independently, many studies have shown that hsp purified from tumors can be used as a tumor-specific vaccine. In this study, we have explored the connection between hsp expression and anti-tumor immunity by transducing murine CT26 colon carcinoma cells with the cDNA of a major hsp, i.e. hsp110. We have shown that over-expression of hsp110 has no effect on CT26 tumor cell growth in vitro, and does not inhibit their anchorage-independent growth capacity. However, in situ, hsp110 over-expressing CT26 tumor (CT26-hsp110) grew at a significantly reduced rate as compared to the wild-type CT26 tumor in immunocompetent mice. Moreover, immunization of mice with inactivated CT26-hsp110 cells significantly inhibited the growth of wild-type CT26 tumor. This immunity was associated with an increased frequency of tumor-specific T cells after vaccination. An in vivo antibody depletion assay demonstrated that inactivated CT26-hsp110 cells elicited anti-tumor responses involving CD8(+) T cells and natural killer (NK) cells, but not CD4(+) T cells. Lastly, the effect of the addition of granulocyte-macrophage colony stimulating factor (GM-CSF) to these vaccine formulations was determined. Mice immunized with irradiated CT26-hsp110 cells combined with GM-CSF-producing bystander cells revealed a complete inhibition of CT26 tumor growth, indicating a synergy between inactivated CT26-hsp110 vaccine activity and GM-CSF. These observations demonstrate that manipulation of hsp110 expression in tumors, specifically when combined with GM-CSF, represents a potentially powerful approach to cancer vaccine formulation.

Targeted Immunotherapy Using Reconstituted Chaperone Complexes of Heat Shock Protein 110 and Melanoma-associated Antigen Gp100

Cancer Research. May, 2003  |  Pubmed ID: 12750279

This report defines a novel approach to heat shock protein vaccine formulation that takes advantage of the chaperoning property of heat shock protein hsp110 to efficiently bind a large protein substrate (specifically, human melanoma-associated antigen gp100) during heat shock. We demonstrate that hsp110 can form chaperone complexes with gp100 and prevent heat-induced aggregation of gp100. The resultant natural hsp110-gp100 complexes are strongly immunogenic as determined by their ability to elicit an antigen-specific IFN-gamma production and a cytotoxic T-cell response. Immunization with the hsp110-gp100 complex protected mice against subsequent challenge with human gp100-transduced B16 melanoma, which involves both CD4(+) and CD8(+) T-cell populations. Administration of the hsp110-gp100 vaccine also significantly suppressed the growth of established tumors in a therapeutic model. Furthermore, the hsp110-gp100 chaperone complex exhibited inhibitory effects on the progression of wild-type B16 tumor, suggesting that the induced immune response by human gp100 cross-reacts with mouse gp100. More importantly, the antitumor response obtained with the hsp110-gp100 complex is more potent than that obtained using Complete Freund's Adjuvant with gp100, whereas no response was observed against mouse hsp110 itself. Thus, the use of hsp110 to form natural chaperone complexes with tumor protein antigens such as gp100 represents a powerful approach to therapeutic vaccine formulation with significant potential for clinical application.

HSP110-HER2/neu Chaperone Complex Vaccine Induces Protective Immunity Against Spontaneous Mammary Tumors in HER-2/neu Transgenic Mice

Journal of Immunology (Baltimore, Md. : 1950). Oct, 2003  |  Pubmed ID: 14530326

Heat shock proteins (HSPs) are shown to be strong immunoadjuvants, eliciting both innate and adaptive immune responses against cancers. HSP110 is related in sequence to HSP70 and is approximately 4-fold more efficient in binding to and stabilizing denatured protein substrates compared with HSP70. In the present study we evaluated the ability of a heat shock complex of HSP110 with the intracellular domain (ICD) of human HER-2/neu to elicit effective antitumor immune responses and to inhibit spontaneous mammary tumors in FVB-neu (FVBN202) transgenic mice. The HSP110-ICD complex was capable of breaking tolerance against the rat neu protein and inhibiting spontaneous mammary tumor development. This vaccine induced ICD-specific IFN-gamma and IL-4 production. Depletion studies revealed that CD8(+) T cells were involved in protection against challenge with mouse mammary tumors, whereas CD4(+) T cells revealed partial protection. Increased IgG2a Ab titer in the sera of tumor-free animals after vaccination and elevated CD4(+) CD25(+) regulatory T cells in the PBL of tumor-bearing animals suggested that IFN-gamma-producing Th1 cells may be responsible for partial protection of CD4(+) T cells against the mammary tumor challenge, whereas CD4(+)CD25(+) regulatory T cells (Th2 cells) may suppress the antitumor immune responses. Together, these results suggest that HSP110-ICD complex can elicit effective IFN-gamma-producing T cells against spontaneous mammary tumors and that up-regulation of CD4(+) CD25(+) regulatory T cells may prevent complete eradication of the tumor following immunotherapy.

Development of Cancer Vaccines Using Autologous and Recombinant High Molecular Weight Stress Proteins

Methods (San Diego, Calif.). Jan, 2004  |  Pubmed ID: 14624871

High molecular weight heat shock proteins (HSPs), hsp110 and grp170, derived from cancer cells have been previously shown to elicit tumor-specific immunity. This phenomenon is attributed to the antigenic peptides associated with the HSPs. Based on the unique chaperoning properties of these HSPs, a new vaccination strategy has been recently developed to elicit antigen-specific antitumor immunity. This approach utilizes tumor-associated antigens naturally complexed to these highly efficient molecular chaperones under heat shock conditions. This chapter focuses on the methodologies of these two vaccine strategies: I. purification of hsp110 and grp170 from tumor tissue or cell lines; II. generation and characterization of in vitro HSP-antigen complexes by heat shock using recombinant HSPs derived from a baculovirus protein expression system.

Cancer Immunotherapy and Heat-shock Proteins: Promises and Challenges

Expert Opinion on Biological Therapy. Mar, 2004  |  Pubmed ID: 15006730

Recent mechanistic studies on the role of heat-shock proteins (HSPs) to induce innate and adaptive immune responses have resulted in conflicting reports. Whereas some groups reported that HSPs have direct immunological function, others emphasised the endotoxin contamination of HSP preparations and questioned the antigen-specificity of HSP vaccines. The present review will discuss these issues and suggest that HSPs have diverse and distinct immunological functions that could be superimposed on effects resulting from endotoxin contamination or misunderstood by using experimental procedures with inadequate controls. To understand the actual function of HSPs in their interaction with the immune system, methods and procedures need to be optimised and appropriate controls need to be used. These points should also clarify the conflicting findings about HSPs and promote our knowledge about other immuologically important components that may be present in HSP preparations.

Heat Shock Proteins and Scavenger Receptors: Role in Adaptive Immune Responses

Immunological Investigations. 2005  |  Pubmed ID: 16136784

Tumor-derived heat shock proteins have shown promise as anti-cancer vaccines in clinical trials. Heat shock proteins (HSPs) can generate potent anti-tumor immunity and elicit antigen-specific CD8+ T cell responses in murine studies. Antigen presenting cells (APC), such as macrophages and dendritic cells (DCs), can elicit antigen-specific CD8+ T cell responses mediated by HSPs. CD91 was the first identified endocytic scavenger receptor for HSPs on APC that can facilitate the process of cross-presentation. Other scavenger receptors may also play a similar role in this process. The present review critically evaluates the identified HSP endocytic receptors on APCs that may generate adaptive immune responses. A better understanding of this interaction between HSPs and APCs may further unravel mechanisms of immunoadjuvant function of HSPs.

HSP110 Induces "danger Signals" Upon Interaction with Antigen Presenting Cells and Mouse Mammary Carcinoma

Immunobiology. 2005  |  Pubmed ID: 16164037

HSP110 is a large molecular weight heat shock protein highly capable of chaperoning large proteins. When chaperoning tumour antigens, HSP110 is capable of eliciting effective anti-tumour immune responses. In the present study, we have determined whether such immunoadjuvant properties of HSP110 stem from its ability to induce "danger signals" through interaction with antigen presenting cells (APCs) and with tumour cells. In the previous studies, endotoxin contamination of HSP preparations was always a matter of concern and controversy. Therefore, we prepared recombinant HSP110 with low endotoxin concentration at which LPS did not have any effect on dendritic cells (DCs). We then evaluated the ability of the HSP110 to induce "danger signals" while interacting with APCs or mouse mammary carcinoma cell line (MMC), as evaluated by modulation of cell surface receptors and cytokines involved in innate and adaptive immune responses. We also performed competition studies in order to rule out contribution of endotoxin in HSP110 preparations while interacting with DCs and MMC. We showed that low endotoxin HSP110 induced DCs to up-regulate the expression of MHC class II, CD40 and CD86 molecules, and to secrete pro-inflammatory cytokines IL-6, IL-12 and TNF-alpha. Importantly, HSP110 induced MMC to secrete IL-12 and elevate secretion of IL-6 and expression of CD40 molecule. These findings demonstrate that HSP110 acts as a "danger signal" through its interaction with DCs and tumour cells, regardless of its endotoxin component. These immunoadjuvant properties of HSP110 suggest that pre-existing immunity in tumour-bearing individuals,may be due to the release of HSPs from tumours upon necrosis alerting the immune system against the tumours.

Emergence of Immune Escape Variant of Mammary Tumors That Has Distinct Proteomic Profile and a Reduced Ability to Induce "danger Signals"

Breast Cancer Research and Treatment. Apr, 2006  |  Pubmed ID: 16211331

Breast tumors are shaped, in part, by a process termed immunoediting which selects for immunologically evasive phenotypes. In the present study we used the rat neu-transgenic mouse model of breast cancer and its congenic non-transgenic parental strain, FVB, to explore the phenotype of tumors that emerge in the presence of an immune response directed against the neu antigen. When inoculated into parental FVB mice, a neu-overexpressing mouse mammary carcinoma (MMC) cell line isolated from spontaneous breast tumors of the FVB neu (FVBN202) transgenic mouse, elicited a neu-specific immune response resulting in a tumor rejection because of the presence of the rat neu antigen. However, a neu negative variant (ANV) of MMC arose after a long latency in spite of the neu-specific immune response. We show that compared to MMC, ANV tumor cells have a significantly reduced ability to secrete pro-inflammatory cytokines and the CCL5 chemokine, to express immunostimulatory chaperones, and they have a distinct expression of proteins involved in cell motility, and metabolic and signal transduction pathways. These studies suggest that tumor escape through immunoediting can not be explained by the loss of a single tumor antigen, but rather by a selection process of a tumor variant that has a reduced ability to induce "danger signals" together with up-regulation of proteins involved in the tumor survival. Based on these findings, we propose to target novel antigens over-expressed in the escape variant of breast tumors to treat primary tumor and to prevent tumor relapse.

Chaperoning Function of Stress Protein Grp170, a Member of the Hsp70 Superfamily, is Responsible for Its Immunoadjuvant Activity

Cancer Research. Jan, 2006  |  Pubmed ID: 16424054

When used as vaccines, tumor-derived stress proteins can elicit antitumor immune responses. For members of the hsp70 superfamily, like grp170, this seems to be due to (a) the chaperoning of antigenic peptide by the stress protein and (b) the binding of the stress protein to receptor(s) on antigen-presenting cells (APC) and subsequent antigen presentation. This suggests that domains exist on the stress protein for each function. In this study, we determine the ability of grp170 and its structural domains to (a) bind to and present melanoma-associated antigen gp100 to the immune system and (b) to bind to receptors on APCs. A direct correlation between chaperone function, binding to APCs in a receptor-like manner, and antitumor immunity was observed. Two mutants that share no common sequence, yet are both effective in their antitumor activities, compete with one another for APC binding. Studies of other members of the hsp70 superfamily, hsp110 and hsp70, or their domain deletion mutants, further confirmed that APC binding segregates with chaperoning function and not sequence. Therefore, these studies suggest that molecular chaperoning is involved in stress protein interactions with APCs, antigen binding, and in eliciting antitumor immunity, thus bridging this ancient function of stress proteins in prokaryotes to their ability to elicit immunity in higher organisms.

Immunoadjuvant Chaperone, GRP170, Induces 'danger Signals' Upon Interaction with Dendritic Cells

Immunology and Cell Biology. Apr, 2006  |  Pubmed ID: 16519738

When chaperoning tumour antigens, glucose-regulated protein 170 (GRP170) is capable of inducing effective antitumour immune responses. In the present study, we determined whether such immunoadjuvant properties of GRP170 also involve the ability to induce 'danger signals' through interaction with APC. We prepared recombinant GRP170 in the baculovirus expression system with low endotoxin concentration at which LPS did not have any effect on dendritic cells (DC). We showed that GRP170 binds DC in a receptor-mediated fashion and induces DC to upregulate the expression of MHC class II, CD86 and CD40 molecules, and to secrete pro-inflammatory cytokines. GRP170 also induced expression of CD40 molecules in a B16F10 cell line, whereas LPS failed to do so. These findings show that GRP170 acts as a danger signal through its interaction with DC, regardless of its endotoxin component.

Comment on "Tumor Progression Can Occur Despite the Induction of Very High Levels of Self/tumor Antigen-specific CD8+ T Cells in Patients with Melanoma"

Journal of Immunology (Baltimore, Md. : 1950). Apr, 2006  |  Pubmed ID: 16585536

Heat Shock Proteins As Vaccine Adjuvants in Infections and Cancer

Drug Discovery Today. Jun, 2006  |  Pubmed ID: 16713905

In addition to maintaining cell homeostasis under physiological and stress conditions, some heat shock proteins (HSPs) are potent inducers of immunity and have been harnessed as vaccine adjuvants targeted to cancers and infections. HSPs are a group of ubiquitous intracellular molecules that function as molecular chaperones in numerous processes, such as protein folding and transport, and are induced under stress conditions, such as fever and radiation. Certain HSPs are potent inducers of innate and antigen-specific immunity. They activate dendritic cells partly through toll-like receptors, activate natural killer cells, increase presentation of antigens to effector cells and augment T-cell and humoral immune responses against their associated antigens. Their roles in priming multiple host defense pathways are being exploited in vaccine development for cancer and infectious diseases.

Heat Shock Proteins HSP70 and GP96: Structural Insights

Cancer Immunology, Immunotherapy : CII. Mar, 2006  |  Pubmed ID: 16032399

Several heat shock proteins (HSPs) act as potent adjuvants for eliciting anti-tumor immunity. HSP-based tumor vaccine strategies have been highly successful in animal models and are undergoing testing in clinical trials. It is generally accepted that HSPs, functioning as chaperones for tumor antigens, elicit tumor-specific adaptive immune responses. HSPs also appear to induce innate immune responses in an antigen-independent fashion. Innate responses generated by HSPs may contribute to anti-tumor immunity. Immunologically active chaperones with anti-tumor activity are referred to as "immunochaperones". Here, we review the studies that address the role of structural domains or regions of the immunochaperones HSP70 and GP96 that may be involved in the induction of adaptive or innate immune responses.

HER-2/neu Antigen Loss and Relapse of Mammary Carcinoma Are Actively Induced by T Cell-mediated Anti-tumor Immune Responses

European Journal of Immunology. Mar, 2007  |  Pubmed ID: 17304628

Induction of tumor-specific immune responses results in the inhibition of tumor development. However, tumors recur because of the tumor immunoediting process that facilitates development of escape mechanisms in tumors. It is not known whether tumor escape is an active process whereby anti-tumor immune responses induce loss or downregulation of the target antigen in the antigen-positive clones. To address this question, we used rat neu-overexpressing mouse mammary carcinoma (MMC) and its relapsed neu antigen-negative variant (ANV). ANV emerged from MMC under pressure from neu-specific T cell responses in vivo. We then cloned residual neu antigen-negative cells from MMC and residual neu antigen-positive cells from ANV. We found marked differences between these neu-negative clones and ANV, demonstrating that the residual neu-negative clones are probably not the origin of ANV. Since initial rejection of MMC was associated with the presence of IFN-gamma-secreting T cells, we treated MMC with IFN-gamma and showed that IFN-gamma could induce downregulation of neu expression in MMC. This appears to be due to methylation of the neu promoter. Together, these data suggest that neu antigen loss is an active process that occurs in primary tumors due to the neu-targeted anti-tumor immune responses.

Come Forth 1E10 Anti-idiotype Vaccine: Delivering the Promise to Immunotherapy of Small Cell Lung Cancer

Cancer Biology & Therapy. Feb, 2007  |  Pubmed ID: 17426436

Tumor Immunoediting and Immunosculpting Pathways to Cancer Progression

Seminars in Cancer Biology. Aug, 2007  |  Pubmed ID: 17662614

Recent studies have suggested that a natural function of the immune system is to respond and destroy aberrant, dysfunctional cells by a process called immunosurveillance. These studies also suggest that the tumors that arise despite immunosurveillance have been immunosculpted by the immune system. The purported abilities of tumors to induce immune tolerance and suppression, the increased pathogenic behavior of the tumor cells following exposure to immune effectors and the loss of immunogenicity (i.e. immunoediting) often observed in advanced stage tumors could be the result of immunosculpting. In some cases, these immunosculpting features may be permanent and irreversible. However, in other cases, reversible epigenetic mechanisms may underlie the immune resistant tumor phenotype. Regardless, these immune-induced alterations could contribute to cancer pathogenesis. Understanding the mechanisms by which tumors evade immunity will be important for disease prevention and therapeutics.

Danger Signals and Nonself Entity of Tumor Antigen Are Both Required for Eliciting Effective Immune Responses Against HER-2/neu Positive Mammary Carcinoma: Implications for Vaccine Design

Cancer Immunology, Immunotherapy : CII. Sep, 2008  |  Pubmed ID: 18278493

Using parental FVB mice and their neu transgenic counterparts, FVBN202, we showed for the first time that dangerous hyperplasia of mammary epithelial cells coincided with breaking immunological tolerance to the neu "self" tumor antigen, though such immune responses failed to prevent formation of spontaneous neu-overexpressing mammary carcinoma (MMC) or reject transplanted MMC in FVBN202 mice. On the other hand, neu-specific immune responses appeared to be effective against MMC in parental FVB mice because of the fact that rat neu protein was seen as "nonself" antigen in these animals and the protein was dangerously overexpressed in MMC. Interestingly, low/intermediate expression of the neu "nonself" protein in tumors induced immune responses but such immune responses failed to reject the tumor in FVB mice. Our results showed that self-nonself (SNS) entity of a tumor antigen or danger signal alone, while may equally induce an antigen-specific immune response, will not warrant the efficacy of immune responses against tumors. On the other hand, entity of antigen in the context of dangerous conditions, i.e. abnormal/dangerous overexpression of the neu nonself protein, will warrant effective anti-tumor immune responses in FVB mice. This unified "danger-SNS" model suggests focusing on identification of naturally processed cryptic or mutated epitopes, which are considered semi-nonself by the host immune system, along with novel dangerous adjuvant in vaccine design.

Signatures Associated with Rejection or Recurrence in HER-2/neu-positive Mammary Tumors

Cancer Research. Apr, 2008  |  Pubmed ID: 18381452

We have previously shown T-cell-mediated rejection of the neu-overexpressing mammary carcinoma cells (MMC) in wild-type FVB mice. However, following rejection of primary tumors, a fraction of animals experienced a recurrence of a neu antigen-negative variant (ANV) of MMC (tumor evasion model) after a long latency period. In the present study, we determined that T cells derived from wild-type FVB mice can specifically recognize MMC by secreting IFN-gamma and can induce apoptosis of MMC in vitro. Neu transgenic (FVBN202) mice develop spontaneous tumors and cannot reject it (tumor tolerance model). To dissect the mechanisms associated with rejection or tolerance of MMC tumors, we compared transcriptional patterns within the tumor microenvironment of MMC undergoing rejection with those that resisted it either because of tumor evasion/antigen loss recurrence (ANV tumors) or because of intrinsic tolerance mechanisms displayed by the transgenic mice. Gene profiling confirmed that immune rejection is primarily mediated through activation of IFN-stimulated genes and T-cell effector mechanisms. The tumor evasion model showed combined activation of Th1 and Th2 with a deviation toward Th2 and humoral immune responses that failed to achieve rejection likely because of lack of target antigen. Interestingly, the tumor tolerance model instead displayed immune suppression pathways through activation of regulatory mechanisms that included in particular the overexpression of interleukin-10 (IL-10), IL-10 receptor, and suppressor of cytokine signaling (SOCS)-1 and SOCS-3. These data provide a road map for the identification of novel biomarkers of immune responsiveness in clinical trials.

Does HER-2/neu Antigen Loss in Metastatic Breast Tumors Occur Under Immune Pressure?

International Journal of Cancer. Journal International Du Cancer. Sep, 2008  |  Pubmed ID: 18566999

Prostate-derived Ets Transcription Factor As a Favorable Prognostic Marker in Ovarian Cancer Patients

International Journal of Cancer. Journal International Du Cancer. Sep, 2008  |  Pubmed ID: 18567002

We have previously shown that ovarian tumors express prostate-derived Ets transcription factor (PDEF). However, the precise role of PDEF in the prognosis of ovarian cancer is unknown. In our study, we report for the first time that expression of PDEF in tumor lesions of patients with ovarian cancer is associated with favorable prognosis. Evaluation of samples from 40 patients with ovarian cancer showed that early stage (IA) and borderline (IIB, III) ovarian tumors expressed higher levels of PDEF mRNA and protein and lower levels of survivin compared to late stage ovarian tumors (IIIC and IV, p < 0.05). Normal ovarian tissues expressed the highest levels of PDEF mRNA and protein when compared to tumor tissues (p < 0.05). A Log-Rank test showed that overall survival of patients with PDEF-positive and survivin-negative ovarian tumors was significantly longer than those with PDEF-negative and survivin-positive tumors (p < 0.01). Forced expression of PDEF in PDEF-negative ovarian tumor cells inhibited tumor cell growth, induced apoptosis, downregulated survivin expression and its promoter activity. Furthermore, treatment of ovarian cancer cells with vitamin D or a selenium compound resulted in re-expression of PDEF, downregulation of survivin, induction of apoptosis and inhibition of tumor cell growth when compared to untreated controls (p < 0.05). Together, these observations showed an inverse correlation between PDEF and survivin expression and suggested that increased PDEF expression along with reduced survivin was associated with prolonged survival of patients with ovarian cancer.

Adoptive Transfer of HER2/neu-specific T Cells Expanded with Alternating Gamma Chain Cytokines Mediate Tumor Regression when Combined with the Depletion of Myeloid-derived Suppressor Cells

Cancer Immunology, Immunotherapy : CII. Jun, 2009  |  Pubmed ID: 18979098

Adoptive immunotherapy (AIT) using ex vivo-expanded HER-2/neu-specific T cells has shown initial promising results against disseminated tumor cells in the bone marrow. However, it has failed to promote objective responses against primary tumors. We report for the first time that alternating gamma chain cytokines (IL-2, IL-7 and IL-15) ex vivo can expand the neu-specific lymphocytes that can kill breast tumors in vitro. However, the anti-tumor efficacy of these neu-specific T cells was compromised by the increased levels of myeloid-derived suppressor cells (MDSC) during the premalignant stage in FVBN202 transgenic mouse model of breast carcinoma. Combination of AIT with the depletion of MDSC, in vivo, resulted in the regression of neu positive primary tumors. Importantly, neu-specific antibody responses were restored only when AIT was combined with the depletion of MDSC. In vitro studies determined that MDSC caused inhibition of T cell proliferation in a contact-dependent manner. Together, these results suggest that combination of AIT with depletion or inhibition of MDSC could lead to the regression of mammary tumors.

Incubation of Antigen-sensitized T Lymphocytes Activated with Bryostatin 1 + Ionomycin in IL-7 + IL-15 Increases Yield of Cells Capable of Inducing Regression of Melanoma Metastases Compared to Culture in IL-2

Cancer Immunology, Immunotherapy : CII. Oct, 2009  |  Pubmed ID: 19198835

Regression of established tumors can be induced by adoptive immunotherapy (AIT) with tumor draining lymph node (DLN) lymphocytes activated with bryostatin and ionomycin (B/I). We hypothesized that B/I-activated T cells cultured in IL-7 + IL-15 might proliferate and survive in culture better than cells cultured in IL-2, and that these cells would have equal or greater anti-tumor activity in vivo. Tumor antigen-sensitized DLN lymphocytes from either wild-type or T cell receptor transgenic mice were harvested, activated with B/I, and expanded in culture with either IL-2, IL-7 + IL-15 or a regimen of alternating cytokines. Cell yields, proliferation, apoptosis, phenotypes, and in vitro responses to tumor antigen were compared for cells grown in different cytokines. These T cells were also tested for anti-tumor activity against melanoma lung metastases established by prior i.v. injection of B16 melanoma cells. IL-7 + IL-15 or alternating cytokines resulted in much faster and prolonged proliferation and much less apoptosis of B/I-activated T cells than culturing the same cells in IL-2. This resulted in approximately tenfold greater yields of viable cells. Culture in IL-7 + IL-15 yielded higher proportions of CD8+ T cells and a higher proportion of cells with a central memory phenotype. Despite this, T cells grown in IL-7 + IL-15 had higher IFN-gamma release responses to tumor antigen than cells grown in IL-2. Adoptive transfer of B/I-activated T cells grown in IL-7 + IL-15 or the alternating regimen had equal or greater efficacy on a "per-cell" basis against melanoma metastases. Activation of tumor antigen-sensitized T cells with B/I and culture in IL-7 + IL-15 is a promising modification of standard regimens for production of T cells for use in adoptive immunotherapy of cancer.

Immune-induced Epithelial to Mesenchymal Transition in Vivo Generates Breast Cancer Stem Cells

Cancer Research. Apr, 2009  |  Pubmed ID: 19276366

The breast cancer stem cell (BCSC) hypotheses suggest that breast cancer is derived from a single tumor-initiating cell with stem-like properties, but the source of these cells is unclear. We previously observed that induction of an immune response against an epithelial breast cancer led in vivo to the T-cell-dependent outgrowth of a tumor, the cells of which had undergone epithelial to mesenchymal transition (EMT). The resulting mesenchymal tumor cells had a CD24(-/lo)CD44(+) phenotype, consistent with BCSCs. In the present study, we found that EMT was induced by CD8 T cells and the resulting tumors had characteristics of BCSCs, including potent tumorigenicity, ability to reestablish an epithelial tumor, and enhanced resistance to drugs and radiation. In contrast to the hierarchal cancer stem cell hypothesis, which suggests that breast cancer arises from the transformation of a resident tissue stem cell, our results show that EMT can produce the BCSC phenotype. These findings have several important implications related to disease progression and relapse.

Gemcitabine Directly Inhibits Myeloid Derived Suppressor Cells in BALB/c Mice Bearing 4T1 Mammary Carcinoma and Augments Expansion of T Cells from Tumor-bearing Mice

International Immunopharmacology. Jul, 2009  |  Pubmed ID: 19336265

Myeloid derived suppressor cells (MDSCs) accumulate in 4T1 mammary carcinoma bearing mice and present a barrier to the success of adoptive immunotherapy (AIT) by suppressing T cell immunity. In this study, we investigated the inhibition of MDSCs by gemcitabine (GEM), a chemotherapy agent that may have favorable immunologic effects. BALB/c mice were inoculated with 4T1 mammary carcinoma cells and treated with GEM either once a week starting 5 days after tumor inoculation (EARLY GEM) or as a single dose at days 20-25 (LATE GEM). Splenic mononuclear cells were isolated, activated in vitro, expanded, and stimulated with tumor antigen. T cells were then used for AIT to treat tumor-bearing mice. EARLY GEM treatment of 4T1 tumor-bearing mice significantly inhibited tumor growth, reduced splenomegaly, and significantly decreased MDSC proportion in the spleen. Support for a direct effect was demonstrated through suppression of MDSCs in spleens, bone marrow, and blood harvested 24 and 48 h after LATE GEM treatment, despite no significant decrease in tumor burden. Interestingly, treatment of tumor-bearing mice with GEM augmented in vitro expansion of splenic T cells and boosted IFN-gamma secretion in response to stimulation by tumor antigen. However, despite GEM-mediated inhibition of MDSC suppression, splenic T cells from mice with advanced tumors were ineffective in vivo against established tumors. This study provides support for direct inhibition of MDSCs and direct reduction of tumor burden by GEM in 4T1 tumor-bearing mice. GEM treatment of mice with advanced tumors improves T cell function and growth in vitro.

Human T Cells Express CD25 and Foxp3 Upon Activation and Exhibit Effector/memory Phenotypes Without Any Regulatory/suppressor Function

Journal of Translational Medicine. 2009  |  Pubmed ID: 19849846

Foxp3 has been suggested to be a standard marker for murine Tregs whereas its role as marker for human Tregs is controversial. While some reports have shown that human Foxp3+ T cells had no regulatory function others have shown their role in the inhibition of T cell proliferation.

Radiofrequency Thermal Ablation of Breast Tumors Combined with Intralesional Administration of IL-7 and IL-15 Augments Anti-tumor Immune Responses and Inhibits Tumor Development and Metastasis

Breast Cancer Research and Treatment. Apr, 2009  |  Pubmed ID: 18425677

Tumor development or recurrence is always a matter of concern following radiofrequency thermal ablation (RFA) of tumors. To determine whether combining RFA with immunologically active cytokines might induce tumor-specific immune responses against mammary carcinoma and inhibit tumor development or metastasis, we evaluated intralesional injection of IL-7 and IL-15 in RFA-treated murine tumors. We used two different breast carcinoma models: neu-overexpressing mouse mammary carcinoma (MMC) in FVBN202 transgenic mouse and 4T1 tumors in Balb/c mouse. MMC tend to relapse even in the presence of neu-specific immune responses, and 4T1 is a weakly immunogenic, aggressive and highly metastatic transplantable tumor. In vivo growth of both of these tumors is also associated with increased numbers of CD11b+Gr1+ myeloid-derived suppressor cells (MDSC). We showed for the first time that unlike RFA alone, RFA combined with the administration of intralesional IL-7 and IL-15 (after RFA), induced immune responses to tumors, inhibited tumor development and lung metastasis, and reduced MDSC.

GM-CSF is One of the Main Breast Tumor-derived Soluble Factors Involved in the Differentiation of CD11b-Gr1- Bone Marrow Progenitor Cells into Myeloid-derived Suppressor Cells

Breast Cancer Research and Treatment. Aug, 2010  |  Pubmed ID: 19898981

Recent reports have shown the involvement of tumor burden as well as GM-CSF in supporting myeloid-derived suppressor cells (MDSC). However, it is not known what progenitor cells may differentiate into MDSC in the presence of GM-CSF, and whether FVBN202 transgenic mouse model of spontaneous breast carcinoma may exhibit distinct subset distribution of CD11b+Gr1+ cells. In addition, it is not known why CD11b+Gr1+ cells derived from tumor-free and tumor-bearing animals exhibit different functions. In this study, we determined that GM-CSF was one of the tumor-derived soluble factors that induced differentiation of CD11b-Gr1- progenitor cells from within monocytic/granulocytic bone marrow cells into CD11b+Gr1+ cells. We also showed that CD11b+Gr1+ cells in FVBN202 mice consisted of CD11b+Ly6G-Ly6C+ suppressive and CD11b+Ly6G+Ly6C+ non-suppressive subsets. Previously reported variations between tumor-free and tumor-bearing animals in the function of their CD11b+Gr1+ cells were found to be due to the variations in the proportion of these two subsets. Therefore, increasing ratios of CD11b+Gr1+ cells derived from tumor-free animals revealed their suppressive activity on T cells, in vitro. Importantly, GM-CSF supported the generation of CD11b+Ly6G-Ly6C+ suppressor subsets that inhibited proliferation as well as anti-tumor function of neu-specific T cells. These findings suggest revisiting the use of GM-CSF for the expansion of dendritic cells, ex vivo, for cell-based immunotherapy or as an adjuvant for vaccines for patients with cancer in whom MDSC play a major role in the suppression of anti-tumor immune responses.

Tumour Secreted Grp170 Chaperones Full-length Protein Substrates and Induces an Adaptive Anti-tumour Immune Response in Vivo

International Journal of Hyperthermia : the Official Journal of European Society for Hyperthermic Oncology, North American Hyperthermia Group. 2010  |  Pubmed ID: 20210603

We employed a grp170-secreting tumour cell system to determine whether tumour cells engineered to secrete grp170 generate an antitumour-specific immune response. Further, we examine the possibility that secreted grp170 can bind to and co-transport out of tumour cells full-length tumour antigens that may play a role in the anti-tumour immune response.

IL-7 + IL-15 Are Superior to IL-2 for the Ex Vivo Expansion of 4T1 Mammary Carcinoma-specific T Cells with Greater Efficacy Against Tumors in Vivo

Breast Cancer Research and Treatment. Jul, 2010  |  Pubmed ID: 19826947

Regression of established tumors can be induced by adoptive immunotherapy (AIT) with tumor draining lymph node (DLN) lymphocytes activated with bryostatin and ionomycin (B/I). Tumor antigen-sensitized DLN lymphocytes from BALB/c mice with 10-day 4T1 mammary carcinomas were harvested, activated with B/I, and expanded in culture with either interleukin-2 (IL-2) or IL-7 + IL-15. Cell yields, proliferation, phenotypes, and in vitro responses to tumor antigen were compared for cells grown in different cytokines. These T cells were also tested for antitumor activity against established 4T1 mammary carcinomas after inoculation of tumor cells subcutaneously (s.c.). IL-7/15 resulted in much faster and more prolonged proliferation of B/I-activated T cells than culturing the same cells in IL-2. This resulted in approximately 5-10-fold greater yields of viable cells. Culture in IL-7/15 yielded higher proportions of CD8(+) T cells and a higher proportion of cells with a central memory phenotype. T cells grown in IL-2 had higher interferon-gamma (IFN-gamma) release responses to tumor antigen than cells grown in IL-7/15. Adoptive transfer of B/I-activated T cells grown in IL-7/15 demonstrated much greater efficacy against 4T1 tumors in vivo. Activation of tumor antigen-sensitized T cells with B/I and culture in IL-7 + IL-15 is a promising modification of standard regimens for production of T cells for use in AIT of cancer.

Revisiting Cancer Immunoediting by Understanding Cancer Immune Complexity

The Journal of Pathology. May, 2011  |  Pubmed ID: 21480229

Since 1909, the cancer immunosurveillance concept has undergone four distinct eras. These include a general acceptance during 1957–1974, an abandonment during 1974–1996, resurrection during 1996–2001 in the form of an elegant theory of tumour immunoediting proposed by Robert Schreiber, and a retreat since 2006. Recently, in the Journal of Pathology, Ciampricotti et al reported an elegant experimental model designed by establishing RAG2−/−/MMTV-NeuT mice. Using this, they demonstrated that the development and metastasis of HER-2/neu-positive spontaneous mammary carcinoma were not altered by the presence or absence of the adaptive immune system. Their fascinating results are a call to revisit controversial reports as to an effective role of the adaptive immune system in tumour inhibition versus tumour promotion or tolerance in the development of spontaneous carcinomas. Ciampricotti and colleagues present a strong case for revising our ideas of cancer immunoediting and appreciating the complexity of the interaction between cancer and the immune system.

Comment on "Cutting Edge: CD8+ T Cell Priming in the Absence of NK Cells Leads to Enhanced Memory Responses"

Journal of Immunology (Baltimore, Md. : 1950). Jun, 2011  |  Pubmed ID: 21597038

Prostate-derived Ets Transcription Factor (PDEF) is a Potential Prognostic Marker in Patients with Prostate Cancer

The Prostate. Aug, 2011  |  Pubmed ID: 21656828

Reduced expression of prostate-derived Ets transcription factor (PDEF) leads to morphologic change as well as increased migration and invasiveness of prostate cancer cells. However, the clinical relevance of PDEF expression and its relationship to anti-apoptotic protein survivin is yet to be determined.

Activated NKT Cells and NK Cells Render T Cells Resistant to Myeloid-derived Suppressor Cells and Result in an Effective Adoptive Cellular Therapy Against Breast Cancer in the FVBN202 Transgenic Mouse

Journal of Immunology (Baltimore, Md. : 1950). Jul, 2011  |  Pubmed ID: 21670315

Attempts to cure breast cancer by adoptive cellular therapy (ACT) have not been successful. This is primarily due to the presence of tumor-induced immune-suppressive mechanisms as well as the failure of tumor-reactive T cells to provide long-term memory responses in vivo. To address these clinically important challenges, we developed an ex vivo protocol for the expansion of tumor-reactive immune cells obtained from tumor-bearing animals prior to or after local radiation therapy. We used an Ag-free protocol that included bryostatin 1/ionomycin and sequential common γ-chain cytokines (IL-7/IL-15 + IL-2). The proposed protocol expanded tumor-reactive T cells as well as activated non-T cells, including NKT cells, NK cells, and IFN-γ-producing killer dendritic cells. Antitumor efficacy of T cells depended on the presence of non-T cells. The effector non-T cells also rendered T cells resistant to myeloid-derived suppressor cells. Radiation therapy altered phenotypic distribution and differentiation of T cells as well as their ability to generate central memory T cells. ACT by means of the expanded cells protected animals from tumor challenge and generated long-term memory responses against the tumor, provided that leukocytes were derived from tumor-bearing animals prior to radiation therapy. The ex vivo protocol was also able to expand HER-2/neu-specific T cells derived from the PBMC of a single patient with breast carcinoma. These data suggest that the proposed ACT protocol should be studied further in breast cancer patients.

CD4+ T Cells Inhibit the Neu-specific CD8+ T-cell Exhaustion During the Priming Phase of Immune Responses Against Breast Cancer

Breast Cancer Research and Treatment. Apr, 2011  |  Pubmed ID: 20480224

Studies conducted in animal model of infectious diseases or H-Y antigen model suggest a crucial role for CD4+ T cells in providing help for CD8+ T-cell memory responses. This concept suggests that inclusion of T helper epitopes in vaccine formulation will result in improved CD8+ T-cell responses. Although this concept has been applied to cancer vaccine design, the role of CD4+ T cells in the memory differentiation of CD8+ T cells and retention of their anti-tumor function have never been tested in breast cancer model. Using the FVB mouse model of neu-positive breast carcinoma we report for the first time that helpless T cells showed cytostatic or tumor inhibitory effects during primary tumor challenge whereas, helped T cells showed cytotoxic effects and resulted in complete tumor rejection. Such differential effects, in vivo, were associated with higher frequency of CD8+PD-L1+ and CD8+PD-1+ T cells in animals harboring helpless T cells as well as higher titer of IL-2 in the sera of animals harboring helped T cells. However, depletion of CD4+ T cells did not alter the ability of neu-specific CD8+ T cells to differentiate into memory cells and to retain their effector function against the tumor during recall challenge. These results suggest the inhibitory role of CD4+ T cells on CD8+ T-cell exhaustion without substantial effects on the differentiation of memory T cells during priming phase of the immune responses against breast cancer.

Adoptive Cell Therapy of Prostate Cancer Using Female Mice-derived T Cells That React with Prostate Antigens

Cancer Immunology, Immunotherapy : CII. Mar, 2011  |  Pubmed ID: 21088965

In this study, we report a novel treatment strategy that could potentially be used to improve efficacy of adoptive cell therapy for patients with prostate cancer. We show that female C57BL/6 mice are able to effectively reject two syngeneic prostate tumors (TRAMP-C2 and RM1) in a T cell-dependent manner. The protective antitumor immunity appears to primarily involve T cell responses reactive against general prostate tumor/tissue antigens, rather than simply to male-specific H-Y antigen. For the first time we show that adoptive transfer of lymphocytes from TRAMP-C2-primed or naïve female mice effectively control prostate tumor growth in male mice, when combined with host pre-conditioning (i.e., non-myeloablative lymphodepletion) and IL-2 administration. No pathological autoimmune response was observed in the treated tumor-bearing male mice. Our studies provide new insights regarding the immune-mediated recognition of male-specific tissue, such as the prostate, and may offer new immunotherapy treatment strategies for advanced prostate cancer.

Prostate-derived Ets Transcription Factor (PDEF) is a Potential Prognostic Marker in Patients with Prostate Cancer

The Prostate. Jan, 2011  |  Pubmed ID: 21229552

BACKGROUND: Reduced expression of prostate-derived Ets transcription factor (PDEF) leads to morphologic change as well as increased migration and invasiveness of prostate cancer cells. However, the clinical relevance of PDEF expression and its relationship to anti-apoptotic protein survivin is yet to be determined. METHODS: Tissue microarrays of 73 prostate carcinomas and their adjacent benign prostate tissue, as well as 50 benign prostates were evaluated for PDEF expression by immunohistochemistry. Results were confirmed in available tumor tissues using Western blot and RT-PCR. Expression of survivin in prostate carcinoma and benign tissues were determined using Western blot. Results and correlation with clinical data were statistically analyzed. RESULTS: Patients' specimens with low Gleason scores (GS < 5) expressed higher levels of PDEF protein and lower levels of survivin protein when compared with moderate-to-high GS tumors (GS > 6). Patients with PDEF-positive tumor survived significantly longer (P < 0.0001) than patients with PDEF-negative tumor, and the 8-year survival rate was 94% and 40%, respectively. PDEF expression was detected at the highest levels in benign tissues and was down-regulated or lost in 30 recently diagnosed prostate carcinomas. Re-expression of PDEF in prostate cancer cells inhibited survivin expression. Treatment of prostate cancer cells with methylseleninic acid resulted in restoration of PDEF expression, down-regulation of survivin, and inhibition of tumor cell growth when compared with untreated controls (P < 0.05). CONCLUSIONS: These studies demonstrated an inverse correlation between PDEF and survivin expression, and that up-regulation of PDEF was associated with a favorable prognosis in patients with clinically localized prostate cancer. Prostate © 2010 Wiley-Liss, Inc.

Tumor Escape and Progression of HER-2/neu Negative Breast Cancer Under Immune Pressure

Journal of Translational Medicine. 2011  |  Pubmed ID: 21453513

Emerging data from pre-clinical and clinical studies suggest that HER-2/neu-specific T cell responses could induce HER-2/neu antigen loss in the tumor cells. These data suggest that patients with HER-2/neu negative breast cancer might have had HER-2/neu positive premalignant lesions in the past that progressed to HER-2/neu negative breast cancer under HER-2/neu-specific immune pressure.

An Immunologic Portrait of Cancer

Journal of Translational Medicine. 2011  |  Pubmed ID: 21875439

The advent of high-throughput technology challenges the traditional histopathological classification of cancer, and proposes new taxonomies derived from global transcriptional patterns. Although most of these molecular re-classifications did not endure the test of time, they provided bulk of new information that can reframe our understanding of human cancer biology. Here, we focus on an immunologic interpretation of cancer that segregates oncogenic processes independent from their tissue derivation into at least two categories of which one bears the footprints of immune activation. Several observations describe a cancer phenotype where the expression of interferon stimulated genes and immune effector mechanisms reflect patterns commonly observed during the inflammatory response against pathogens, which leads to elimination of infected cells. As these signatures are observed in growing cancers, they are not sufficient to entirely clear the organism of neoplastic cells but they sustain, as in chronic infections, a self-perpetuating inflammatory process. Yet, several studies determined an association between this inflammatory status and a favorable natural history of the disease or a better responsiveness to cancer immune therapy. Moreover, these signatures overlap with those observed during immune-mediated cancer rejection and, more broadly, immune-mediated tissue-specific destruction in other immune pathologies. Thus, a discussion concerning this cancer phenotype is warranted as it remains unknown why it occurs in immune competent hosts. It also remains uncertain whether a genetically determined response of the host to its own cancer, the genetic makeup of the neoplastic process or a combination of both drives the inflammatory process. Here we reflect on commonalities and discrepancies among studies and on the genetic or somatic conditions that may cause this schism in cancer behavior.

Targeting the Immunoregulator SRA/CD204 Potentiates Specific Dendritic Cell Vaccine-induced T-cell Response and Antitumor Immunity

Cancer Research. Nov, 2011  |  Pubmed ID: 21914786

Although dendritic cell (DC) vaccines offer promise as cancer immunotherapy, further improvements are needed to amplify their clinical therapeutic efficacy. The pattern recognition scavenger receptor SRA/CD204 attenuates the ability of DCs to activate CD8(+) T-cell responses. Therefore, we examined the impact of SRA/CD204 on antitumor responses generated by DC vaccines and we also evaluated the feasibility of enhancing DC vaccine potency by SRA/CD204 blockade. DCs from SRA/CD204-deficient mice were more immunogenic in generating antitumor responses to B16 melanoma, compared with DCs from wild-type mice. Similarly, siRNA-mediated knockdown of SRA/CD204 by lentiviral vectors improved the ability of wild-type DCs to stimulate the expansion and activation of CD8(+) T cells specific for idealized or established melanoma antigens in mice. Using SRA/CD204-silenced DCs to generate antigen-targeted vaccines, we documented a marked increase in the level of antitumor immunity achieved against established B16 tumors and metastases. This increase was associated with enhanced activation of antigen specific CTLs, greater tumor infiltration by CD8(+) T cells and NK cells, and increased intratumoral ratios of both CD4(+) and CD8(+) T-effector cells to CD4(+)CD25(+) T-regulatory cells. Our studies establish that downregulating SRA/CD204 strongly enhances DC-mediated antitumor immunity. In addition, they provide a rationale to enhance DC vaccine potency through SRA/CD204-targeting approaches that can improve clinical outcomes in cancer treatment.

Distinct Signatures of the Immune Responses in Low Risk Versus High Risk Neuroblastoma

Journal of Translational Medicine. 2011  |  Pubmed ID: 21978632

Over 90% of low risk (LR) neuroblastoma patients survive whereas less than 30% of high risk (HR) patients are long term survivors. Age (children younger than 18 months old) is associated with LR disease. Considering that adaptive immune system is well developed in older children, and that T cells were shown to be involved in tumor escape and progression of cancers, we sought to determine whether HR patients may tend to show a signature of adaptive immune responses compared to LR patients who tend to have diminished T-cell responses but an intact innate immune response.

Favorable Outcomes in Patients with High Donor-Derived T Cell Count After In Vivo T Cell-Depleted Reduced-Intensity Allogeneic Stem Cell Transplantation

Biology of Blood and Marrow Transplantation : Journal of the American Society for Blood and Marrow Transplantation. Oct, 2011  |  Pubmed ID: 22005648

Patients with hematologic malignancies were conditioned using a rabbit antithymocyte globulin-based reduced-intensity conditioning regimen for allogeneic stem cell transplantation. Donor-derived CD3(+) cell count (ddCD3), a product of CD3(+) cell chimerism and absolute CD3(+) cell count, when <110/μL at 8 weeks post-stem cell transplantation predicted a high risk of sustained mixed chimerism and relapse. Alternatively, patients with a higher ddCD3 developed graft-versus-host disease more frequently, and when partially chimeric, had higher rates of conversion to full donor chimerism after withdrawal of immunosuppression. Early data from our small cohort of patients indicate that ddCD3 at 8 weeks may be used to guide decisions regarding withdrawal of immunosuppression and administration of donor lymphocyte infusion in partially T cell-depleted reduced-intensity regimens.

Suppression of Antigen-specific CD4(+) T Cell Activation by SRA/CD204 Through Reducing the Immunostimulatory Capability of Antigen-presenting Cell

Journal of Molecular Medicine (Berlin, Germany). Nov, 2011  |  Pubmed ID: 22083206

Pattern recognition scavenger receptor SRA/CD204, primarily expressed on specialized antigen-presenting cells (APCs), including dendritic cells (DCs) and macrophages, has been implicated in multiple physiological and pathological processes, including atherosclerosis, Alzheimer's disease, endotoxic shock, host defense, and cancer development. SRA/CD204 was also recently shown to function as an attenuator of vaccine response and antitumor immunity. Here, we, for the first time, report that SRA/CD204 knockout (SRA(-/-)) mice developed a more robust CD4(+) T cell response than wild-type mice after ovalbumin immunization. Splenic DCs from the immunized SRA(-/-) mice were much more efficient than those from WT mice in stimulating naïve OT-II cells, indicating that the suppressive activity of SRA/CD204 is mediated by DCs. Strikingly, antigen-exposed SRA(-/-) DCs with or without lipopolysaccharide treatment exhibited increased T-cell-stimulating activity in vitro, which was independent of the classical endocytic property of the SRA/CD204. Additionally, absence of SRA/CD204 resulted in significantly elevated IL12p35 expression in DCs upon CD40 ligation plus interferon gamma (IFN-γ) stimulation. Molecular studies reveal that SRA/CD204 inhibited the activation of STAT1, mitogen activated protein kinase p38, and nuclear factor-kappa B signaling activation in DCs treated with anti-CD40 antibodies and IFN-γ. Furthermore, splenocytes from the generated SRA(-/-) OT-II mice showed heightened proliferation upon stimulation with OVA protein or MHC-II-restricted OVA(323-339) peptide compared with cells from the SRA(+/+) OT-II mice. These results not only establish a new role of SRA/CD204 in limiting the intrinsic immunogenicity of APCs and CD4(+) T cell activation but also provide additional insights into the molecular mechanisms involved in the immune suppression by this molecule.

A Signature of Immune Function Genes Associated with Recurrence-free Survival in Breast Cancer Patients

Breast Cancer Research and Treatment. Feb, 2012  |  Pubmed ID: 21479927

The clinical significance of tumor-infiltrating immune cells has been reported in a variety of human carcinomas including breast cancer. However, molecular signature of tumor-infiltrating immune cells and their prognostic value in breast cancer patients remain elusive. We hypothesized that a distinct network of immune function genes at the tumor site can predict a low risk versus high risk of distant relapse in breast cancer patients regardless of the status of ER, PR, or HER-2/neu in their tumors. We conducted retrospective studies in a diverse cohort of breast cancer patients with a 1-5 year tumor relapse versus those with up to 7 years relapse-free survival. The RNAs were extracted from the frozen tumor specimens at the time of diagnosis and subjected to microarray analysis and real-time RT-PCR. Paraffin-embedded tissues were also subjected to immunohistochemistry staining. We determined that a network of immune function genes involved in B cell development, interferon signaling associated with allograft rejection and autoimmune reaction, antigen presentation pathway, and cross talk between adaptive and innate immune responses were exclusively upregulated in patients with relapse-free survival. Among the 299 genes, five genes which included B cell response genes were found to predict with >85% accuracy relapse-free survival. Real-time RT-PCR confirmed the 5-gene prognostic signature that was distinct from an FDA-cleared 70-gene signature of MammaPrint panel and from the Oncotype DX recurrence score assay panel. These data suggest that neoadjuvant immunotherapy in patients with high risk of relapse may reduce tumor recurrence by inducing the immune function genes.

CD44(+)/CD24(-/low) Cancer Stem/progenitor Cells Are More Abundant in Triple-negative Invasive Breast Carcinoma Phenotype and Are Associated with Poor Outcome

Human Pathology. Mar, 2012  |  Pubmed ID: 21835433

Women classified as having triple-negative tumors have a poor prognosis. The importance of CD44(+)/CD24(-/low) (stem/progenitor cell-phenotype) in breast cancer patients has also been appreciated. However, correlation between triple negativity and CD44(+)/CD24(-/low) with tumor recurrence remains elusive. In the present study, we evaluated tumor specimens of 50 breast cancer patients with known hormone receptor status for whom we had follow-up information and outcome data available, and performed immunohistochemistry analysis to determine CD44 and CD24 expression. Gene expression arrays were also independently performed on 52 breast cancer specimens with banked frozen tissue. Lastly, we used FVBN202 transgenic mouse model of breast carcinoma and determined the hormone receptor status, the proportion of CD44(+)/CD24(-/low) breast cancer stem-like cells, and the behavior of the tumor. We determined that patients with triple-negative tumors had significantly higher incidence of recurrence or distant metastasis associated with increased frequency of breast cancer stem cell phenotypes compared with those with non-triple-negative tumors. Preclinical studies in FVBN202 transgenic mice confirmed these findings by showing that relapsed tumors were triple negative and had significantly higher frequency of breast cancer stem cells compared with their related primary tumors. Unlike non-triple-negative primary tumors, relapsed triple-negative tumors were tumorigenic at low doses when inoculated into FVBN202 transgenic mice. These findings suggest that CD44(+)/CD24(-/low) breast cancer stem-like cells play an important role in the clinical behavior of triple-negative breast cancer and that development of therapeutic targets directed to breast cancer stem-like cells may lead to reduction in the aggressiveness of triple-negative breast cancers.